The distribution of the cellular retinoic acid-binding protein (CRABP) in some rat tissues has been determined, and the protein has been localized by immunocytochemical techniques in sections from rat testis. In the testis CRABP was found in the seminiferous tubuli with Sertoli cells and the spermatogonia most intensely stained. All other cells of the germinal epithelium appeared largely devoid of CRABP. By use of an enzyme-linked immunosorbent assay CRABP was quantitatively estimated in several tissues and the highest levels were found in testis and eye. Comparisons of the tissue levels of CRABP and of the cellular retinol-binding protein (CRBP) did not reveal any apparent correlation.
The function of vitamin A at the molecular level is these proteins in tissues and cells. In view of the considunknown apart from its well-established role in the vi-erable structural homology that exists between the sual process (Wald, 1968). However, the dependence of known members of this protein superfamily, to which many cell types on the vitamin to maintain normal the retinoid-binding proteins belong, it is difficult to function is well-established. Keratinization and abnor-ascertain that polyclonal antisera react exclusively with ma1 differentiation of several epithelia are well-recog-the appropriate antigen. To circumvent this problem we nized signs of vitamin A deficiency. Other deficiency have raised antisera against synthetic peptides corresymptoms include blindness, increased susceptibility to sponding to sequence stretches unique to CRABP. In infections, retarded growth, and sterility (Moore, 1967). this study we document the reactivity of these antisera Supplementation of retinol (vitamin A alcohol)-deficient and report the quantities of CRABP in some tissues of animals with retinoic acid, the acid derivative of the normal and retinol-deficient, retinoic-acid-fed rats. In vitamin, will alleviate all symptoms of the deficiency addition, the localization of CRABP in sections of testis except the impaired vision and the failure to reproduce have been determined using immunocytochemical (Dowling and Wald, 1960;Thompson et al., 1964). Thus, techniques.
Methods
the metabolic effects of retinol and retinoic acid may be Materials accounted for by separate mechanisms.
To transport the hydrophobic vitamin in the aqueous extra-and intracellular milieu of the body, several pro-Microtiter plates (Nunc immunoplates No. 1) were teins with high affinity for vitamin A compounds have from NUNC (Copenhagen, Denmark). Bovine serum alevolved. The transport of retinol from the storage site in bumin, alkaline phosphatase, and p-nitrophenylphosthe liver to the vitamin A-requiring target cells is ac-phate tablets were from Sigma Chemicals (St. Louis, complished by the plasma retinol-binding protein (RBP; MO). Freund's complete and incomplete adjuvant were for a review see Goodman, 1984). This protein delivers from Difco (Detroit, MI). The reagents used in the aviretinol to target cells by a process involving a specific dine-biotin complex (ABC)-stainings (ABComplex) were interaction of RBP with a cell surface receptor (Rask from DAKO (Copenhagen, Denmark). and Peterson, 1976;Heller, 1975). In several tissues sen-Protein and peptides sitive to retinol deprivation, two intracellular vitamin A-binding proteins have been identified (Chytil and Ong, Rat testis CRABP was purified essentially as de-1984). These proteins, CRBP and CRABP, bind retinol scribed previously (Ong and Chytil, 1978a). Bovine and retinoic acid, respectively, with high specificity. The CRABP was obtained from adrenal glands using essenbinding proteins are structurally related to members of a protein superfamily (Sundelin et al.